Detection of Functional VH81X Heavy Chains in Adult Mice with an Assessment of Complementarity-Determining Region 3 Diversity and Capacity to Form Pre-B Cell Receptor

Abstract

Recent reports have indicated that up to 50% of all H chain proteins formed cannot associate with the surrogate L chain complex and therefore fail to form a pre-B cell receptor (pBCR), which is required for allelic exclusion and, in most cases, verifies that the H chain can assemble with the L chain to form an Ab molecule. Certain VH genes, such as VH81X, appear to be particularly prone to encoding for nonpairing (dysfunctional) H chains. It has been suggested that sequence variability at complementarity-determining region 3, especially when increased by the enzyme TdT, often precludes the ability of VH81X-using H chains to form pBCR. To determine whether a motif exists that accounts for the ability of H chains to pair with surrogate L chain complex/L chain, we have bred a mouse line in which H chain recombination can only occur on one allele, allowing us to compile a pool of H chains capable of forming Ab molecules in the absence of dysfunctional H chains. Somewhat unexpectedly, we have found VH81X H chains capable of Ab formation and cell surface expression in the presence of TdT. Scrutiny of these H chains has revealed that, although highly prone to encode for dysfunctional H chains, sequence variability is not severely limited among functional VH81X H chains. We also demonstrate that surface Ig expression is highly indicative of the capacity of a H chain to form pBCR.

Selection of Ig μ Heavy Chains by Complementarity-Determining Region 3 Length and Amino Acid Composition

“…Our conclusions differ slightly from those of Hayden et al (56), perhaps due to the approach taken by the authors. While their studies looked at the characteristics of V H 81X-HCs that used a number of different J H gene segments (i.e., J H 1 through J H 4), we have focused on V H 81X rearrangements using J H 4.…”

“…In fact, if one looks closely at the data presented by Hayden et al (56), selection biases become apparent. For example, in one group 70% of the sequences use histidine at position 1, whereas only 15% of the sequences use glutamine at this position.…”

Pre-B Cell Receptor Assesses the Quality of IgH Chains and Tunes the Pre-B Cell Repertoire by Delivering Differential Signals

“…It is notable that eight clones, including three of the four, use the same V H 7183.1b and J H 4 segments but carry different CDR3 sequences and differ in their levels of pre-BCR expression (Table I). This finding suggested the importance of the CDR3 region in determining the levels of pre-BCR expression, at least in case of V H 7183.1b-bearing H chains, extending the previous findings (14,37,38). The variability in pre-BCR-forming capacity among H clones was also observed in those using the V H J558 family (Table I) and, hence, appeared not restricted to those using the V H 7183 family even though the average of pre-BCR-forming capacity might depend on the V H segment used.…”

Selection of stereotyped VH81X-μH chains via pre-B cell receptor early in ontogeny and their conservation in adults by marginal zone B cells

“…An intriguing model has been proposed in that pre-B cells expressing SL-non-pairing lH chains have the proliferative advantage over those expressing SL-pairing lH chains in fetal liver, opposed to what happens in the bone marrow (21). Some studies indicated the importance of the particular CDR3 sequences in the selection of V H 81X-lH chains (20,22), whereas another study could not define any sequence constraint in functional V H 81X-lH chains (23). Thus, the molecular mechanism underlying the selection of V H 81X-lH chains during B cell development remains to be clarified.…”